This invention relates to transferring liquids from one container to another, and more particularly the invention relates to methods and apparatus for transferring small quantities of liquids from a multiplicity of depots to a multiplicity of receptacles.
Continuing rapid advances in chemistry, particularly in biochemistry and molecular biology, demand improved capabilities for carrying out large numbers of reactions using small quantities of materials.
In screening patients for genetic disease and susceptibility, for example, the number of conditions for which associated mutations are known is growing, and the numbers of mutant alleles known to be associated with these conditions is increasing. An adequate genetic screen for one or even a few of these conditions can require testing a sample from the patient against a very large number of genetic probes.
Enormous and rapidly increasing numbers of critical biomolecules have been identified and characterized, and an understanding of their various roles in cellular processes is vastly improving. Consequently, for example, the number of potential targets for pharmacologic intervention is very large. Techniques for parallel chemical synthesis, such as combinatorial chemistries, can efficiently produce libraries of large numbers of synthetic compounds that may be screened against selected targets in a rational drug design approach.
Considerable effort has been directed to developing better approaches to handling large numbers of samples, reagents and analytes. Automated laboratory workstations and robotics-based systems have been brought to routine use for some chemical manipulations in screening and synthesis, and dedicated computer applications have been developed both for controlling processes and for manipulating data. And a number of approaches have been proposed for miniaturizing systems for carrying out chemical processes, to reduce the quantities of the various components. Some of these approaches have found use. Particularly, for example, array technologies for binding pair assays use components immobilized in arrays of features on a surface; and microfluidics technologies employ networks of interconnected capillaries to move and combine components on a very small scale.
There is significant and growing interest in employing array technologies for conducting biomolecular manipulations. In array techniques certain of the components are immobilized in a pattern of array features on a surface of a solid support, and permitted to interact with other components. Arrays of binding agents, in which such binding agents as oligonucleotides or peptides are deposited onto a support surface in the form of an array or pattern, can be useful in a variety of applications, including gene expression analysis, drug screening, nucleic acid sequencing, mutation analysis, and the like. For example, information about the nucleotide sequence of a target nucleic acid may be obtained by contacting the target with an array of different surface-bound DNA probes under conditions that favor hybridization of nucleic acids having complementary sequences, and determining at what sites on the array nucleic acid duplexes are formed. Hybridization to surface-bound DNA probe arrays can provide a relatively large amount of information in a single experiment. And, for example, array technology can be useful in differential gene expression analysis.
Such arrays may be prepared in any of a variety of different ways. For example, DNA arrays may be prepared manually by spotting DNA onto the surface of a substrate with a micro pipette. See, Khrapko et al. (1991), DNA Sequence 1:375-388. Or, a dot-blot approach or a slot-blot approach may be employed in which a vacuum manifold transfers aqueous DNA samples from a plurality of wells to a substrate surface. Or, an array of pins can be dipped into an array of fluid samples and then contacted with the substrate surface to produce the array of sample materials. Or, an array of capillaries can be used to produce biopolymeric arrays, as described for example in International Patent Publication WO 95/35505.
U.S. patent applications Ser. Nos. 09/150,504 and 09/150,507 describe forming biomolecular arrays by adaptations of devices employed in the printing industry and, particularly, of inkjet print heads and of automated devices for moving a print head over a print surface and for depositing the inks at desired locations on the surface. These references and others cited herein, above and below, are incorporated herein in their entirety by reference. Other uses of inkjet printing devices to dispense biochemical agents such as proteins and nucleic acids are suggested or disclosed in, for example, U.S. Pat. Nos. 5,658,802; 5,338,688; 5,700,637; 5,474,796; 4,877,745; and 5,449,754.
Whether the miniaturized system is a microfluidic device or an array, or is of some other design, at least some of the various biomolecules to be introduced to the system are typically prepared in depots remote from the receptacles by which they are introduced to the system. These depots may take the form of a multiwell plate (conventionally providing 96 wells in a 12xc3x978 format), for example, or a microtiter plate (conventionally providing 384 wells in a 16xc3x9724 format, or 1536 wells in a 32xc3x9748 format). A technical challenge is presented by the step of transferring the liquids containing the various biomolecules from the depots to the specific receptacles. In an array system constructed using an inkjet printing technique, for example, a technical challenge is presented by the need to transfer the liquids from the depots to the specific reservoirs in the print head.
Conventionally a pipet may be employed to transfer a liquid dropwise from a depot to a receptacle (such as a reservoir in a microfluidics device or a reservoir in a print head). The tip of the pipet is first dipped into the liquid in the depot and some of the liquid is drawn into the pipet; then the pipet is moved to the receptacle and a quantity of the liquid is expelled into the receptacle. Several pipets may be ganged and used to transfer several different liquids at once, to reduce the number of repetitions, but problems of small dimension may make such an approach impractical. In any event the transfer step results in contamination of pipets, which accordingly must be either discarded and replaced or decontaminated (for example by rinsing) before they are used to transfer different liquids. Where a large number of different liquids are to be moved, the transfer apparatus becomes mechanically unwieldy, and the cost of minimizing the risk of contamination is increased.
In one general aspect the invention features a method for transferring liquids from a plurality of wells to one or more receptacles, by displacing liquid contained in each well so that a convex meniscus swells from the opening of the well, and contacting a receptacle with the swollen meniscus to draw at least a portion of the liquid into the receptacle. According to the invention, the liquid transfer is effected directly from the depots to the corresponding receptacles without contact between depots and the receptacles, and without interposition of any transfer device between depots and the receptacles. And, according to the invention, the flow of the liquid into the receptacle following contact of the receptacle with the meniscus is at least initially a result of capillary interaction, and ordinarily is principally so.
In preferred embodiments the wells are arranged in a specified pattern in a depot member, so that where different wells contain different liquids or contain liquids containing different constituents, the different liquid contents are associable with the positions of the wells in the pattern. And, in some embodiments where liquids from different wells are to be transferred to specified different receptacles, the receptacles are arranged in a corresponding or complementary pattern, so that a specified plurality of the wells may be aligned with a corresponding specified plurality of the receptacles, so that transfer of the liquids may be accomplished all at once from the specified wells to the specified receptacles.
Accordingly in another aspect the invention features a method for transferring liquids from a plurality of wells having openings arranged in a selected format to a plurality of receptacles arranged in a corresponding or complementary format, by displacing the liquid contained in each well so that a convex meniscus swells from the opening, and contacting the corresponding receptacle with the swollen meniscus to draw a portion of the liquid into the receptacles.
In some embodiments the liquid displacing step is carried out by inwardly deforming a wall of each well to displace the liquid; in preferred embodiments the wall is inwardly deformed by application of mechanical or fluid pressure to the wall. In other embodiments the liquid displacing step is carried out by introducing a gas into a part of each well away from the opening; in preferred embodiments the gas is introduced through a vent in a part of the wall away from the opening, and in some embodiments the gas is passed through a gas-permeable membrane covering the vent.
In some embodiments the arrangements of the well openings and the receptacles is such that receptacles to which transfer of liquid is specified may come into contact with swollen menisci at the openings of specified wells. In some embodiments the arrangement of either the well openings or the receptacles is in a generally planar format, and the step of contacting the receptacles with the menisci is carried out by bringing the specified receptacles with the menisci at the specified well openings. Or, the arrangement of the well openings and the arrangement of receptacles each is in a generally planar format, and the step of contacting the receptacles with the menisci is carried out by bringing the well openings into respective planes into generally parallel proximity.
In another general aspect the invention features apparatus for transferring a plurality of liquids, the apparatus including a depot member having a plurality of wells each having an inwardly deformable wall portion and an opening, in which the openings are supported in a selected format, and a member defining a plurality of receptacles in a corresponding or complementary format; means for displacing liquid contained within the wells toward and through the openings; and means for bringing well openings and receptacles into proximity. According to the invention, transfer of liquid is effected by deploying the displacing means to displace the liquid in the well, causing a convex meniscus to swell outward from the opening. When a receptacle which has been brought into proximity contacts the swollen meniscus, the liquid is drawn into the receptacle. The apparatus for effecting the transfer is uncomplicated and can be made in a straightforward manner from inexpensive materials using simple tools.
It can for some processes be advantageous to transfer a multiplicity of liquids from a multiplicity of specified wells or depots to a multiplicity of assigned or specified receptacles in a single transfer operation. Accordingly in some embodiments the well openings and the receptacles are arranged so that a multiplicity of corresponding or complementary receptacles and wall openings can be brought into proximity simultaneously, so that the receptacles contact the respective menisci at nearly the same time. Where the well openings are arranged in a generally planar pattern, for example, liquid droplets expressed at a line of such wells may in one step be transferred into a line of receptacles that are brought into generally parallel proximity with the line of wall openings; or, liquid droplets at a planar group of such wells may in one step be transferred into a complementary group of receptacles, themselves arranged in a generally planar pattern, that are brought into generally parallel proximity with the group of well openings.
Accordingly, in some embodiments the receptacle-defining member is generally planar, and the well openings are supported in a generally planar format. In some embodiments the receptacle-defining member is an orifice plate of a print head and the receptacles are in fluid communication with reservoirs in the print head; and in some embodiments the print orifices are the receptacles.
In some embodiments the wells include a deformable wall portion, and the means for displacing the liquid in the wells include means for inwardly deforming the deformable wall portion. In some embodiments the wall-deforming means includes mechanical means such as a plunger for pressing against an outer surface of the deformable wall portion; or means for applying fluid pressure (liquid or gas) at the outer surface of the deformable wall portion.
In other embodiments each well includes a vent positioned away from the opening, and means for introducing a fluid (gas or liquid) through the vent and into the well, to displace the liquid in the well toward the opening. In preferred embodiments the vent is covered by a membrane that retains the liquid in the well under operating conditions, but is permeable to the fluid to be introduced through the vent into the well to displace the liquid in the well.
In some embodiments the well includes a rigid wall portion in addition to the deformable wall portion, and in some embodiments the inwardly deformable wall portion and the rigid wall portion are formed of a unitary piece of material. In some embodiments the inwardly deformable wall portion includes a plastic or elastic film. In some embodiments the inwardly deformable wall portion and the support for the well openings are formed of a unitary piece of material.
Other aspects and advantages of the invention will become apparent from the following detailed description, taken together with the accompanying drawings, illustrating by way of example the principles of the invention.